*2.4. Construction and Preparation of Recombinant DNA*

HSP23-encoding genes of different kinetoplastid species including *L. donovani* (LdBPK\_340230), *L. major* (LmjF.34.0210), *L. infantum* (LinJ.34.0230), *L. braziliensis* (LbrM.20.0220), *Trypanosoma brucei* (Tb927.10.2620), were amplified from species-specific genomic DNA using primer pairs that introduce a *Kpn*I and a *Bcl*I or *BamH*I (for *L. infantum* only) restriction sites (Table S1). Fragments were subsequently ligated into the *Leishmania* expression plasmid pCL1S [42] previously digested with *Kpn*I and *Bgl*II.
